Enzyme-linked Immunoabsorbent Assay Research Articles

Enhancing Retinoic Acid-mediated Effects Through Inhibition of CYP26A1, CYP26B1 and HGF Signaling in Neuroblastoma Cells.

Retinoic acid (RA) induces tumor cell differentiation in diseases like acute promyelocytic leukemia or high-risk neuroblastoma. However, the formation of resistant cells, which results from dysregulation of different signaling pathways, limits therapy success. The present study aimed to characterize basic regulatory processes induced by the application of RA in human neuroblastoma cells, to identify therapeutic targets independent of the often amplified oncogene MYCN. In MYCN-amplified Kelly and MYCN non-amplified SH-SY5Y cells, different assays were employed to quantify the viability and cytotoxicity, while RA-mediated expression changes were examined using genome-wide gene expression analysis followed by quantitative PCR. Enzyme-linked immunoabsorbent assays (ELISA) and western blots were used to determine the levels or activation of the examined proteins. In Kelly cells, treatment with 5 μM RA for 3 days significantly reduced the cell number due to attenuated proliferation, while SH-SY5Y cells were less responsive. An up-regulation of the RA-metabolizing enzymes CYP26A1 and CYP26B1 was observed in both cell lines, and co-treatment with the selective CYP26 inhibitor talarozole markedly decreased cell viability. When RA and ketoconazole, which inhibits CYP26 as well as RA-degrading CYP3A enzymes, were co-administered, not only cell survival was impaired in both cell lines, but also the release of hepatocyte growth factor (HGF). Accordingly, co-application of the c-Met inhibitor tepotinib and RA or ketoconazole substantially decreased cell viability. Independent of MYCN amplification, inhibitors of RA metabolism or HGF signaling might prevent the emergence of RA-resistant neuroblastoma cells when co-applied with RA.

Association between changes in serum bone metabolism markers and bone microarchitecture changes during basic combat training – The ARMI study

ImportanceU.S. Army Basic Combat Training (BCT) improves tibial volumetric bone mineral density (BMD) and structure in most, but not all soldiers. Few studies have investigated whether changes in serum bone biomarkers during BCT are associated with changes in tibial BMD and bone structure following BCT. ObjectiveTo characterize bone biomarker changes during BCT and to investigate the relationship between changes in bone biomarkers and changes in tibial BMD and bone structure. MethodsWe enrolled 235 trainees entering BCT in this ten-week prospective observational study. Trainees provided fasted blood samples and questionnaires weekly throughout BCT. Procollagen type 1 N-terminal propeptide (PINP) and C-terminal telopeptide of type 1 collagen (CTX) were measured by enzyme-linked immunoabsorbent assays every two weeks during BCT. We evaluated body composition and mass via dual-energy X-ray absorptiometry and bone structure, microarchitecture, and mineral density at the distal tibia via high-resolution peripheral quantitative computed tomography at baseline and post-BCT. ResultsBoth male (n = 110) and female trainees (n = 125) were young (20.9 ± 3.7 and 20.7 ± 4.3 years, respectively), with normal to overweight BMIs (25.2 ± 4.1 and 24.2 ± 3.6 kg/m2, respectively). In female trainees, PINP increased during and post-BCT compared to baseline, with the greatest increase in PINP at week four (45.4 % ± 49.6, p < 0.0001), whereas there were no changes in CTX. PINP also increased in male trainees, but only at weeks two and four (21.9 % ± 24.5, p = 0.0027 and 35.9 % ± 35.8, p < 0.0001, respectively). Unlike female trainees, in males, CTX was lower than baseline at weeks four, eight, and post-BCT. The change in PINP from baseline to week four of BCT was positively associated with changes in tibial BMD, Tb.BMD, Tb.Th, Tb.BV/TV, Ct.Th, Ct.Ar, and Ct.Po from the baseline to post-BCT. ConclusionThe bone formation marker PINP increases during U.S. Army BCT, especially during the first four weeks. Increases in PINP, but not CTX, were correlated with improved BMD and bone structure in the distal tibia.

Impact of Some Immunological Parameters (antioxidant – cytokines) in Cutaneous leishmaniasis in a Sample of Patients in the Al-Ramadi City.

Background: Cutaneous leishmaniasis is one of the endemic diseases in Iraq. Both types of cutaneous leishmaniasis that cause different ulcers are present, namely Leishmania major and Leishmania tropica, The disease also represents a major public and global health problem all over the world. Objective: Investigation of the relationship between parasitic infection with cutaneous leishmaniasis and some antioxidants Superoxide, Nitric oxide and immune cytokines tumor necrosis factor-alpha, Interferon-gamma. Materials and Methods: The demographic and epidemiological study included 55 cases suspected of being infected with cutaneous leishmaniasis. They visited Ramadi Teaching Hospital, with 25 volunteers. After obtaining their consent, they were used as control samples, where blood serum samples were obtained from both study groups. After that, work was done in laboratories. Department of Biology - College of Education for Pure Sciences - Anbar University. Results: The patients and healthy people were of different genders, and among the 55 samples from the infected, the number of males was 36 (65.5)% and females 19 (34.5)%, with statistically significant differences p&lt;0.05, and the age groups of the patients were from one year to 60 years. With an average age of (31 years), they were distributed into 6 groups. Also, among the 25 samples from the control group, the number of males was 12 (48%) and females were 13 (52%), Their ages also ranged from one year to 60 years, with an average age of (13) years, the enzyme-linked immunoabsorbent assay (ELISA) technique was used to detect the level of immunological indicators TNF-α, IFN-γ, SOD, NO, where the results showed High levels in the serum of cutaneous leishmaniasis patients (120.00 ± 52.11, 123.00 ± 44.21, 3.10 ± 1.21 and 121.93 ± 33.22), respectively compared to healthy subjects (72.86 ± 14.94, 21.91 ± 45.40, 0.88 ± 0.12, and 81.46 ± 4.50 ) respectively With statistically significant differences at the level of p &lt; 0.05, p &lt; 0.001. Conclusions: The inflammatory cytokines TNF-α, IFN-γ play a vital role in increasing or decreasing the severity in the pathogenesis of the disease and eliminating it.

Bioactive peptides in preterm human milk: Impact of maternal characteristics and their association to neonatal outcomes.

Human milk adipokines in term babies seem partially determined by maternal factors and affect infant's development. We aimed to describe bioactive peptide concentration in very preterm human milk and associations to maternal characteristics and postnatal growth. Mothers delivering ≤32 weeks of gestation and their infant/s were recruited. At 4 weeks of lactation, an aliquot of 24-h-pooled milk was collected for exclusively breastfeeding dyads. Insulin, leptin, adiponectin, and milk fat globule epidermal growth factor-8 (MFG-E8) were measured by enzyme-linked immunoabsorbent assay in skimmed milk. One hundred mothers (28.8 ± 2.3 weeks at delivery) provided a milk sample. Milk insulin was related to gestational age, pre-pregnancy body mass index (BMI), and galactagogue treatment (final model: adjusted R2 : 0.330, p < 0.0001; adjusted β coefficients: galactagogue treatment: 0.348, p 0.001; pre-pregnancy BMI: 0.274, p 0.009; gestational age: -0.290, p 0.007). Adiponectin was higher in mothers with gestational diabetes (30.7 ± 6.5 vs. 24.8 ± 8 ng/mL, p 0.044). Leptin was associated with pre-pregnancy BMI (Spearman's ρ: 0.648, p < 0.0001) and MFG-E8 to presence of labor and multiple pregnancy (final linear regression model, R2 : 0.073, p 0.028, adjusted β coefficients: presence of labor -0.229, p 0.050; twins: -0.192, p 0.099). Milk adiponectin was associated with a greater decrease in length z-scores from birth to 28 days (Pearson's r: -0.225, p 0.032) and to discharge (Pearson's r: -0.290, p 0.003). Milk MFG-E8 was lower in milk of mothers whose babies experienced late-onset sepsis (13.3 ± 5.8 vs. 16.8 ± 6.3 μg/mL, p 0.023). Adipokines levels in preterm human milk are partially related to maternal metabolic status. Milk peptide concentration associates with early neonatal growth trajectories.

Therapeutic Drug Monitoring of Anti-TNFα Inhibitors: A Matter of Cut-Off Ranges.

Therapeutic drug monitoring (TDM) is a useful tool for optimising the use of anti-TNFα inhibitors in patients with inflammatory bowel diseases (IBDs). Recently, point-of-care methods for the quantification of drug levels and anti-drug antibodies (ADAs) have been developed to overcome the limitations of conventional enzyme-linked immunoabsorbent assays (ELISAs). Here, we evaluated the performance, interchangeability, and agreement between an automated ELISA-based immunoassay (CHORUS Promonitor) and the lateral flow assay (RIDA®QUICK) for the quantification of infliximab (IFX, n = 65) and adalimumab (ADM, n = 58) plasma levels in IBD patients. Thirty-two samples for IFX and twenty-three samples for ADM that tested positively for the presence of ADAs were also used. Overall, data analysis showed a good agreement of ADM trough concentrations (R2 = 0.75) between the two assays as well as for ADA measurement (K > 0.8). However, IFX levels highlighted a weak correlation (R2 = 0.58) between the two kits, with the RIDA®QUICK assay overestimating IFX plasma values by 30% when compared to the CHORUS Promonitor kit. Results from this study show that the two assays are not quantitatively and qualitatively interchangeable due to substantial discrepancies in some results. Accordingly, the same assay should be used for the longitudinal follow-up of IBD patients.

Interleukin 1α and interleukin 18 in patients with vitiligo – Results of a case-control study

IntroductionVitiligo is a depigmenting disorder resulting from loss of functional melanocytes in the skin. Variety of inflammatory mediators participate in the regulation of melanogenesis in melanocytes: interleukin-18 (IL-18), interleukin-33, granulocyte-macrophage colony stimulating factor, interferon-γ, prostaglandin E2 have the effect of promoting melanogenesis, while interleukin-1 (IL-1), interleukin-4, interleukin-6, interleukin-17 and tumor necrosis factor can inhibit melanogenesis. AimEvaluation of IL-1α and IL-18 levels in peripheral blood in patients with vitiligo compared to healthy controls. Materials and methodsFifty patients aged 18–81 with vitiligo participated in the study. The control group consisted of 38 healthy people. Venous blood samples were obtained from each participant. Serum IL-1α and IL-18 concentrations were determined using the enzyme-linked immunoabsorbent assay (ELISA). ResultsAmong patients with vitiligo, the mean concentration of IL-1α was 0.13 (± 0.535) pg/mL, while in the control group it was 0.51 (± 1.51). There were no statistically significant differences in IL-1α concentrations between patients in the study group compared to the control group (p > 0.05). In the study group, the mean IL-18 concentration was 141.05 (± 136.33) pg/mL vs 137.33 (± 105.83) pg/mL in the controls. There were no statistically significant differences in IL-18 concentrations between patients in the study group compared to the controls (p > 0.05). In the Spearman correlation test, no correlation was confirmed between IL1α and IL-18 concentrations in the group of patients with vitiligo vs healthy people. ConclusionsThere is no correlation between Il-1 and Il-18 concentration in the blood sera of patients with vitiligo.

Role of Inflammatory/Immune Response and Cytokine Polymorphisms in the Severity of Chronic Hepatitis C (CHC) before and after Direct Acting Antiviral (DAAs) Treatment.

Host regulatory immune response is involved in the hepatic inflammatory process caused by the hepatitis C virus (HCV). We aimed to determine if HCV clearance with direct-acting antivirals (DAAs) changes the hepatic fibrosis stage, biochemical parameters of liver injury, and inflammatory/immune responses. Sample: 329 chronic hepatitis C (CHC) patients, 134 of them treated with DAAs. Liver fibrosis was evaluated by transient elastography (FibroScan), biochemical and cellular parameters were determined by standard methods, cytokine concentration by enzyme-linked immunoabsorbent assay (ELISA), and genetic polymorphisms by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) or endpoint genotyping. Before DAA treatment, severe fibrosis or cirrhosis (F3/4) was associated with higher values of tumor necrosis factor-alpha (TNF-α) and genotypes transforming growth factor-beta-509 C/T_CC (TGF-β-509 C/T_CC), interleukine-10-1082 T/C_CC (IL-10-1082 T/C_CC), and IL-10-592 G/T_GT. After DAA treatment, fewer F3/4 patients and lower values of TNF-α were found. Patients with TNF-α-308 G/A_GG and IL-10-592 G/T_GT were at risk for F3/4. Lack of improvement of liver fibrosis was associated with lower baseline values of platelet count for genotypes TNF-α-308 G/A_GG and haplotype TT/GG of IL-10-1082 T/C and IL-10-592 G/T. Our study showed decreased liver fibrosis/inflammation and normalization of liver injury biomarkers after DAA treatment. It also points to the importance of suppressing the pro-inflammatory response by DAAs in the resolution of hepatitis C, contributing to the improvement of liver damage evaluated by transient elastography.

The relationship between inflammatory markers and vitamin D levels with the severity of coronary artery disease in elderly patients

Introduction: Coronary artery disease (CAD) is the accumulation of atherosclerotic plaques in the coronary arteries. CAD is the most common cause of death worldwide, and its prevalence continues to increase, especially in the elderly population. Atherosclerosis is a chronic inflammatory condition that involves various inflammatory markers, including tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6). In the inflammatory process, vitamin D plays an essential role in developing atherosclerotic plaques affecting the severity of CAD. The elderly are susceptible to vitamin D deficiency and an increase in the inflammatory process related to the severity of CAD. Therefore, this study aims to prove the relationship between inflammatory markers and vitamin D levels with the severity of CAD in elderly patients. Methods: This study used a cross-sectional design in stable CAD patients aged 60 - 75 years who had passed through coronary arteriography. The severity was assessed from the angiography results using the Synergy between PCI with Taxus and Cardiac Surgery (SYNTAX) score, which 3 cardiovascular consultants evaluated and the average outcome was calculated. TNF-α, IL-6, and vitamin D were taken during angiography and examined using high sensitivity enzyme-linked immunoabsorbent assay (ELISA). The correlation test was analyzed using Spearman's Rank correlation test. Results: A total of 38 subjects were included in this study, where 30 (78.9%) patients were male. The mean age of the patients was 64.79 years, and the prevalence of vitamin D deficiency was 57.9%. There was a moderately significant positive correlation between markers of inflammation TNF-α (r=0.499, p=0.001) and IL-6 (r=0.518, p=0.001) and the SYNTAX score. A weak negative correlation was also discovered between vitamin D and the SYNTAX score (r=-0.335, p=0.040). Conclusion: The levels of inflammatory markers (TNF-α and IL-6) and vitamin D correlated with the severity of CAD in elderly patients.

Seroprevalence of infectious bronchitis virus and avian reovirus in free backyard chickens.

Infectious bronchitis virus (IBV) and avian reovirus (ARV) cause significant losses in the poultry industry throughout the world. A cross-sectional study was conducted in four villages in Manjacaze district, Southern Mozambique, to determine the seroprevalence of IBV and ARV. A total of 467 serum samples from adult unvaccinated backyard chickens were screened using commercial and competitive enzyme-linked immunoabsorbent assay kits. Our results showed anti-IBV and anti-ARV antibodies in all surveyed households and villages. The overall seroprevalence was 89.5% (95% confidence interval [CI]: 77.2-97.4) and 95.7% (95% CI: 88.0-99.2) for IBV and ARV, respectively. The risk of becoming exposed to IBV was lower in Chidenguele village compared with the other three villages (p 0.05). However, no statistically significant differences were observed for becoming exposed to ARV between villages (p 0.05). The backyard chickens tested in this study had no previous history of vaccination, outbreaks or typical clinical signs of IB and AR diseases. Therefore, the presence of antibodies to IBV and ARV was considered clear evidence that the birds have been naturally exposed to those two infectious agents, and the infection was of subclinical type. It is concluded that IBV and ARV are widespread in backyard chickens in the studied area. These obtained data are essential for design and implementation of chicken health development programmes.Contribution:The epidemiology of IBV and ARV of backyard chicken in Mozambique is unknown. This study determined the seroprevalence of IBV and ARV in backyard chicken health. The obtained data are essential for design and implementation of chicken health development programmes.

A case-control study on immunologic markers of patients with vitiligo

Vitiligo is a depigmentation skin disease often coexisting with other autoimmune disorders. The prevalence is 0.5–2 % of the world's population. A combination of genetic, environmental and biochemical factors play a role in pathogenesis of the disease. The aim of the study was to measure selected cytokines in the blood sera of patients with vitiligo and to select the best marker of the disease. The study was conducted on 50 patients with vitiligo and 38 controls. The type of vitiligo, body surface area (BSA), disease advancement assessment according to Vitiligo European Task Force, the degree of skin involvement and degree of progression were measured. Patients' skin phototype was determined according to Fitzpatrick’s classification. Medical history was recorded. Venous blood samples were obtained, The sera were used for laboratory test. Determinations of IL-17, IL-22 and IL-23 were performed using the enzyme-linked immunoabsorbent assay. In the study group 4 cases had phototype I, 38 II, 8 III. In the control group: 3, 29, 6, respectively. Universal vitiligo was found in 38 patients, segmental in 2, acro-facial in 10. In all cases VETF was + 1. Hypothyroidism was recorded in the medical histories of 11 cases vs 2 controls. Significantly higher concentrations of IL-22 and IL-23 were in cases vs controls. IL-22 concentrations were significantly higher in the study group with BSA ≤ 10 than in the control group, and in the group with BSA ≥ 10 they were the highest (p < 0.05). IL-22 is the best marker of active universal type vitiligo, it is directly proportional to the extent of lesions.

The Effect of Ginseng Extract on Serum Interleukin-6 Levels in Patients with Community-Acquired Pneumonia

Community-Acquired Pneumonia (CAP) is the most common cause of death and illness in the world. Increased IL-6 can be used as an early indicator of infection or inflammation. Ginseng is a popular herbal medicine. The anti-inflammatory effect of Ginseng is mediated by its ability to inhibit Nuclear Factor Kappa Beta (NF-kB), a proinflammatory regulator to initiate the synthesis of cytokines TNF-α, IL-1β, IL-6, and IL-8. Clinical trial research, quasi-experimental design with a pretest-posttest approach was carried out on 26 community pneumonia patients who were hospitalized at Dr. Moewardi Hospital, Surakarta from October 2020 to January 2021 using purposive sampling. The independent variable was Ginseng extract (GinsanaR) at a dose of 2x100 mg and the dependent variable was serum IL-6 levels. Serum IL-6 levels were measured using the Sandwich Enzyme-Linked Immunoabsorbent Assay (ELISA) method. Mean IL-6 levels in the control group on day 0, day 3, and day 14 were 232,89+156,61 pg/mL, 113,46±83.30 pg/mL and 66.18±66.02 pg/mL, respectively (p=&lt;0.001). Mean IL-6 levels in the treatment group on day 0, day 3, and day 14 were 519,55±609,19 pg/mL, 205.41±329.17 pg/mL and 133,59±291,68 pg/mL, respectively (p=&lt;0.001). Delta IL-6 levels in the control group and the treatment group on day 3 compared to day 0, the mean of the IL-6 control group -119,42±111,70 pg/mL, the mean for the IL-6 treatment group -314,14±532,16 pg/mL; On day 14 compared to day 0, the mean of the IL-6 control group was -166,70±135,54 pg/mL, the mean of the IL-6 treatment group was -385,96±547,10 pg/mL; On day 14 compared to day 3, the mean IL-6 control group was -47.28±47.47, the mean IL-6 control group was -71.82±58.16. The post hoc test (Wilcoxon) obtained a p-value &lt; 0.05, suggesting that Ginseng extract has a significant effect on reducing serum IL-6 serum levels in community pneumonia patients.

Association between Leptin (G2548A) and Leptin Receptor (Q223R) Polymorphisms with Plasma Leptin, BMI, Stress, Sleep and Eating Patterns among the Multiethnic Young Malaysian Adult Population from a Healthcare University.

Relative leptin resistance in childhood to absolute leptin resistance in maturity suggests sleep, eating behaviour, and the psychological state as probable causes. The current body of research provides inconclusive evidence linking G2548A and Q223R to obesity. Furthermore, we could find very little data that have observed the association between the environment and gene polymorphism, especially in the multiethnic population that exists in Malaysia. This study searched for a possible link between sleeping habits, eating behaviour, and stress indicators with plasma leptin and its genetic variation in young adult Malaysian healthcare students. The study involved 185 first- and second-year medical and dental students from a healthcare university. Polymerase Chain Reaction–Restriction Fragment Length Polymorphism(PCR-RFLP) determined the genotype, Enzyme Linked Immunoabsorbant Assay (ELISA) tested the serum leptin, and a self-administered questionnaire evaluated sleep, eating behaviour, and psychological condition. Gender and ethnicity are linked to fasting plasma leptin levels (p < 0.001). Plasma leptin also affects stress, anxiety, and sadness. Leptin (LEP) and Leptin Receptor (LEPR) polymorphisms were not associated with BMI, plasma leptin, sleep, eating behaviour, or psychological state. Young adult Malaysian Indians were obese and overweight, while Chinese were underweight. These findings imply overweight and obese participants were in stage I of leptin resistance and lifestyle change or leptin therapy could prevent them from becoming cripplingly obese as they age.

Advanced Glycation End Product (AGE) and Soluble Receptor of AGE (sRAGE) Levels in Relation to Periodontitis Severity and as Putative 3-Year Outcome Predictors in Patients Undergoing Coronary Artery Bypass Grafting (CABG).

Tissue concentrations of advanced glycation end product (AGE) and peripheral soluble receptor of AGE (sRAGE) levels may be associated with periodontitis severity. Both parameters and periodontitis might serve as outcome predictors for patients undergoing coronary artery bypass grafting (CABG). This study aimed to investigate possible associations between periodontitis and AGE/sRAGE. Ultimately, we wanted to examine whether AGE, sRAGE, and severe periodontitis are associated with the incidence of new cardiovascular events within 3 years of follow-up after CABG. Ninety-five patients with coronary vascular disease (CVD) (age 69 years, 88.3% males) needing CABG surgery were included. Periodontal diagnosis was made according to the guidelines of the “Centers for Disease Control and Prevention (CDC)” (2007) and staged according to the new classification of periodontal diseases (2018). AGE tissue concentrations were assessed as skin autofluorescence (sAF). sRAGE levels were determined by using a commercially available enzyme-linked immunoabsorbance assay (ELISA) kit. Univariate and multivariate baseline and survival analyses were carried out with Mann–Whitney U test, Chi² test, Kaplan–Meier curves with Log-Rank test, and logistic and Cox regression. sAF was identified as an independent risk indicator for severe periodontitis with respect to the cofactors age, gender, plaque index, and diabetes (adjusted odds ratio [OR] = 2.9, p = 0.028). The degree of subgingival inflammation assessed as a percentage of sites with bleeding on probing (BOP) was inversely correlated with sRAGE concentration (r = −0.189, p = 0.034). Both sAF (Hazard Ratio [HR] = 2.4, p = 0.004) and sRAGE (HR = 1.9, p = 0.031) increased the crude risk for new adverse events after CABG. The occurrence of severe periodontitis trends towards a higher risk for new cardiovascular events (HR = 1.8, p = 0.115). Applying multivariate Cox regression, only peripheral arterial disease (adjusted HR = 2.7, p = 0.006) and history of myocardial infarction (adjusted HR = 2.8, p = 0.010) proved to be independent risk factors for cardiovascular outcome. We conclude that sAF may represent a new, independent risk indicator for severe periodontitis. In contrast, sAF, sRAGE, and severe periodontitis were not independent prognostic factors for postoperative outcome in patients undergoing CABG.

TRAUMA-DERIVED EXTRACELLULAR VESICLES ARE SUFFICIENT TO INDUCE ENDOTHELIAL DYSFUNCTION AND COAGULOPATHY.

Although a number of studies have demonstrated increased release of extracellular vesicles (EVs) and changes in their origin differentials after trauma, the biologic significance of EVs is not well understood. We hypothesized that EVs released after trauma/hemorrhagic shock (HS) contribute to endotheliopathy and coagulopathy. To test this hypothesis, adoptive transfer experiments were performed to determine whether EVs derived from severely injured patients in shock were sufficient to induce endothelial dysfunction and coagulopathy. Methods: Total EVs were enriched from plasma of severely injured trauma/HS patients or minimally injured patients by ultracentrifugation and characterized for size and numbers. Under isoflurane anesthesia, noninjured naive C57BL/6J mice were administered EVs at varying concentrations and compared with mice receiving equal volume vehicle (phosphate-buffered saline (PBS)) or to mice receiving EVs from minimally injured patients. Thirty minutes after injection, mice were sacrificed, and blood was collected for thrombin generation (thrombin-antithrombin, thrombin-antithrombin complex [TAT] assay) and syndecan-1 by enzyme-linked immunoabsorbent assay (ELISA). Lungs were harvested for examination of histopathologic injury and costained with von Willebrand factor and fibrin to identify intravascular coagulation. Bronchial alveolar lavage fluid was aspirated from lungs for protein measurement as an indicator of the endothelial permeability. Data are presented as mean ± SD, P < 0.05 was considered significant, and t test was used. Results: An initial proof-of-concept experiment was performed in naive mice receiving EVs purified from severely injured trauma/HS patients (Injury Severity Score [ISS], 34 ± 7) at different concentrations (5 × 106 to 3.1 × 109/100 μL/mouse) and compared with PBS (control) mice. Neither TAT nor syndecan-1 levels were significantly different between groups at 30 minutes after EV infusion. However, lung vascular permeability and histopathologic injury were significantly higher in the EV group, and lung tissues demonstrated intravascular fibrin deposition. Based on these data, EVs from severely injured trauma/HS patients (ISS, 32 ± 6) or EVs from minimally injured patients (ISS, 8 ± 3) were administered to naive mice at higher concentrations (1 × 109 to 1 × 1010 EV/100 μL/mouse). Compared with mice receiving EVs from minimally injured patients, plasma TAT and syndecan-1 levels were significantly higher in the trauma/HS EV group. Similarly, bronchial alveolar lavage protein and lung histopathologic injury were higher in the trauma/HS EV group, and lung tissues demonstrated enhanced intravascular fibrin deposition. Conclusion: These data demonstrate that trauma/HS results in the systemic release of EVs, which are capable of inducing endotheliopathy as demonstrated by elevated syndecan-1 and increased permeability and coagulopathy as demonstrated by increased TAT and intravascular fibrin deposition. Targeting trauma-induced EVs may represent a novel therapeutic strategy.

Evaluation of the Thermal Stability of a Vaccine Prototype Based on Virus-like Particle Formulated HIV-1 Envelope.

The long-term storage stability of vaccines has a major impact on the roll-out and success of global immunization programs. For the Human Immunodeficiency Virus type 1 (HIV-1) virus-like particle (VLP) vaccine prototype evaluated here, nanoparticle tracking analysis (NTA), and enzyme-linked immunoabsorbent assay (ELISA) results demonstrated a remarkable structural stability. VLPs maintained their integrity and the recognition of relevant B-cell epitopes for three months at 4 and −20 °C. Interestingly, most particles remained intact and preserved the recognition of relevant epitopes even after a week of storage at room temperature.

Comparative analyses in transcriptome of human granulosa cells and follicular fluid micro-environment between poor ovarian responders with conventional controlled ovarian or mild ovarian stimulations

BackgroundBoth mild and conventional controlled ovarian stimulation are the frequently used protocols for poor ovarian responders. However, there are some debates about which treatment is better. Moreover, little is known about the follicular physiology after the two ovarian stimulation protocols. This study was intended to investigate the features in granulosa cells and follicular fluid micro-environment after the two different ovarian stimulation protocols in poor responders.MethodsGranulosa cells RNA were sequenced using Illumina Hiseq technology. Specific differently expressed genes and proteins were verified by real-time quantitative PCR and Western blot analysis. Moreover, hormone and cytokine concentrations in the follicular fluid were measured by electrochemiluminescence immunoassay and enzyme-linked immunoabsorbent assay. The correlation between the results of molecular experiments and the laboratory outcomes were analyzed by Spearman correlation analysis.ResultsThe differentially expressed genes between the two groups were involved in 4 signaling pathways related to the follicular development; three proteins pertinent to the TGF-β signaling pathway were expressed differently in granulosa cells between the two, and the constituents in the follicular fluid were also different. Further, a correlation between the TGF-β signaling pathway and the good-quality embryo was observed.ConclusionsThe present study made a comparison for the first time in the transcriptome of human granulosa cells and the follicular fluid micro-environment between poor responders with the conventional controlled ovarian stimulation or the mild ovarian stimulation, showing that the TGF-β signaling pathway may correlate with the good-quality of embryos in the mild group, which may be instrumental to the choice of optimal management for IVF patients.

Detection of aflatoxins in tomato and pepper pastes in Istanbul, Turkey

The aim of this study is to determine the amount of total aflatoxins and aflatoxin B1 (AFB1) in tomato and pepper pastes sold in farmer’s markets in Istanbul to evaluate whether aflatoxin levels were within the standards regarded as safe (<5 µg kg-1). For this purpose, a total of 64 samples, including 26 tomato pastes, 15 paprika pastes, and 23 chili pastes, were analysed using two different detection methods, enzyme-linked immunoabsorbent assay (ELISA) and high-performance liquid chromatography (HPLC). By ELISA, aflatoxins were detected in 27 samples (1–2.5 µg kg-1), and AFB1 was detected at a level of 1 µg kg-1 in 20 samples. Furthermore, 21 of 64 samples were found to contain aflatoxins (0.21–2.34 µg kg-1), with 16 of them contaminated with AFB1 (0.22–2.34 µg kg-1) by HPLC. The obtained result showed that both methods are suitable for aflatoxin determination in tomato and pepper paste samples and the samples have been proven to be within the standards considered as safe.

Irisin response to acute moderate intensity exercise and high intensity interval training in youth of different obesity statuses: A randomized crossover trial.

Limited data exist regarding the impact of an acute bout of exercise with varying intensities on irisin levels in the youth of different obesity statuses. The objectives were to (1) compare an acute bout of moderate continuous intensity (MCI) exercise and an acute bout of high‐intensity interval training (HIIT) on irisin response in youth with different obesity statuses and, (2) investigate whether changes in irisin levels are correlated with exploratory outcomes. A randomized crossover design study was conducted on 25 youth aged 12–18 years old. Participants were classified as either healthy weight (BMI percentile <85; n = 14) or overweight/obese (BMI percentile ≥85; n = 11). Participants performed an MCI exercise session at 50% of heart rate reserve for 35 min and a HIIT exercise session for 35 min, with intervals every 5 min increasing from 50% heart rate reserve to 85–90% for 2 min. Irisin was measured using an enzyme‐linked immunoabsorbent assay from plasma sampling obtained throughout the exercise (at times 0, 7, 14, 21, 28, and 35 min). A time effect was observed throughout the HIIT session [F(1,5) = 6.478, p < 0.001]. Bonferonni post‐hoc analysis revealed significant differences in irisin levels post‐exercise (35 min) compared to times 7, 14, 21, and 28 min. Irisin increased during HIIT (81.0% ± 71.3; p = 0.012) in youth with a healthy weight. No differences were observed for youth living as overweight or with obesity. Overall, HIIT elicits a higher peak irisin response compared to MCI exercise training in youth.

Concentration of dentin sialoprotein at the initial stage of orthodontic treatment using self-ligating and conventional preadjusted brackets: A pilot study.

OBJECTIVES:This study evaluated differences in concentration of dentin sialoprotein (DSP) in gingival crevicular fluid (GCF) relating to orthodontically induced inflammatory root resorption (OIIRR) at the initial stage of orthodontic treatment using self-ligating and conventional preadjusted brackets.MATERIALS AND METHODS:Eighteen patients were assigned to three groups of equal size. Two experimental groups received non-extraction orthodontic treatment using passive self-ligating or conventional preadjusted bracket. The control group included patients without orthodontic treatment. GCF was collected from five proximal sites of maxillary anterior teeth at subsequent intervals: immediately prior to orthodontic treatment (T0), and at three and 12 weeks after initiation of treatment (T1 and T2). DSP concentration was evaluated by enzyme-linked immunoabsorbent assay and the differences in DSP levels were analyzed between and within groups.RESULTS:There were no significant differences in DSP levels within both experimental groups and the control group during T0-T1-T2 (P ≥ 0.05). A significant difference of DSP concentration was found between the conventional preadjusted bracket and the control group at T2 (P = 0.038). However, it was thought to be clinically insignificant.CONCLUSION:The study showed no significant difference in DSP concentration at the initial stage of orthodontic treatment with either self-ligating or conventional preadjusted bracket.

Human cytomegalovirus autophagy is related to the interferon synthesis and mTOR signal pathway

Introduction: Human cytomegalovirus (HCMV) is reported to be involved in the occurrence of many human diseases. To further investigate the biological changes of HCMV, we analyzed the relevant factors that affect the autophagy caused by HCMV infection. Methods: Firstly, we cultured human embryonic lung fibroblasts (HELF) cells with HCMV infection, and evaluated the effects of HELF cells infected with different viruses through Enzyme-linked immunoabsorbent assay (ELISA), Real-time quantitative Polymerase Chain Reaction (RT-qPCR), Acridine orange (AO) staining and Western blotting (WB) experiments. Results: Through the above experiments, we found that the combined treatment of HCMV infection and carbamazepine, rapamycin and si-mTOR promoted the increase of interferon (IFN)-α/β expression and protein level, and caused the increase of LC3B protein level in HELF cells. In addition, HCMV infection could also affect the biological activities of HELF cells by regulating signal pathways like the JAK/STAT. Conclusion: Autophagy induced by HCMV is affected by the changes in the biological behavior of HELF cells, especially IFN-α/β synthesis and mTOR signal pathway. These findings might shed new light on HCMV-related disease treatment.